Estudo de imidas naftálicas e derivados como ferramentas para a pesquisa de dano oxidativo em Trypanosoma cruzi

Abstract

Chagas Disease, caused by the protozoan Trypanosoma cruzi, afflicts millions world-wide, especially in Latin America. T. cruzi has many strains, which can be divided in two main groups: I and II. Group I is related to the sylvan cycle and group II to the domestic cycle of the parasite (which involves humans). Group I strains present higher resistance to oxidative damage caused by substances like hydrogen peroxide than those from group II. Naphthalic imides are small molecules also capable of generating reactive oxygen species (ROS) but only when stimulated by a specific wavelength radiation (340 nm). Thus, it was our objective to test the effect of two naphthalic imides in a few T. cruzi group I and II representative strains, in order to evaluate the use of such substances as alternative tools to the study of oxidative lesions in that protozoan. We observed that the tested imides were capable of reducing the survival of the oxidative damage sensitive strains (group II) more than the resistant ones (group I), in agreement with previous data for hydrogen peroxide. We also observed that even under conditions where the adequate radiation for activation of the imides wasnt present they would still reduce the survival of the tested strains. Knowing T. cruzi has bioluminescence, we measured that emission and verified it to occur above 300 nm, thus being able to activate the imides at high concentrations. In low concentrations the naphthalic imides can only be activated by lamps of an adequate power (300 to 400 W). We also used a vector peptide sequence consisting of seven arginine residues that, according to the literature, are capable of carrying covalently bonded molecules inside many types of cells. Unfortunately the results were inconclusive. As the product of the MSH2 gene is related to the repair of damaged DNA in many organisms, including T. cruzi, we tested the imides against a knock-out strain for that gene. Its survival was lower when compared to the wild type, indicating lesion in the parasites DNA. As ROS can modify the guanine nucleotide in 8-oxo-7,8-dihydroguanine (8oxoG), we tested the imides in a mutant strain overexpressing the DNA polimerase kappa, which is known to synthesize beyond such modifications. Its survival was higher when compared to the wild type, indicating the imides were modifying guanine into 8oxoG. Concluding, we believe that at low concentrations (40 M) the imides can be used as tools to study oxidative DNA lesions in Trypanosoma cruzi.